biomimetics Article Three-Dimensional Printed Antimicrobial Objects of Polylactic Acid (PLA)-Silver Nanoparticle Nanocomposite Filaments Produced by an In-Situ Reduction Reactive Melt Mixing Process Nectarios Vidakis 1, Markos Petousis 1,* , Emmanouel Velidakis 1, Marco Liebscher 2 and Lazaros Tzounis 3 1 Mechanical Engineering Department, Hellenic Mediterranean University, Estavromenos, 71004 Heraklion, Crete, Greece; [email protected] (N.V.); [email protected] (E.V.) 2 Institute of Construction Materials, Technische Universität Dresden, DE-01062 Dresden, Germany; [email protected] 3 Department of Materials Science and Engineering, University of Ioannina, 45110 Ioannina, Greece; [email protected] * Correspondence: [email protected]; Tel.: +30-2810-37-9227 Received: 14 August 2020; Accepted: 31 August 2020; Published: 2 September 2020 Abstract: In this study, an industrially scalable method is reported for the fabrication of polylactic acid (PLA)/silver nanoparticle (AgNP) nanocomposite filaments by an in-situ reduction reactive melt mixing method. The PLA/AgNP nanocomposite filaments have been produced initially + reducing silver ions (Ag ) arising from silver nitrate (AgNO3) precursor mixed in the polymer melt to elemental silver (Ag0) nanoparticles, utilizing polyethylene glycol (PEG) or polyvinyl pyrrolidone (PVP), respectively, as macromolecular blend compound reducing agents. PEG and PVP were added at various concentrations, to the PLA matrix. The PLA/AgNP filaments have been used to manufacture 3D printed antimicrobial (AM) parts by Fused Filament Fabrication (FFF). The 3D printed PLA/AgNP parts exhibited significant AM properties examined by the reduction in Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) bacteria viability (%) experiments at 30, 60, and 120 min duration of contact (p < 0.05; p-value (p): probability). It could be envisaged that the 3D printed parts manufactured and tested herein mimic nature’s mechanism against bacteria and in terms of antimicrobial properties, contact angle for their anti-adhesive behavior and mechanical properties could create new avenues for the next generation of low-cost and on-demand additive manufacturing produced personal protective equipment (PPE) as well as healthcare and nosocomial antimicrobial equipment. Keywords: Fused Filament Fabrication (FFF); 3D printing; reactive melt processing; antimicrobial (AM) properties; polylactic acid (PLA); silver (Ag) nanoparticles (NPs); polyethylene glycol (PEG); polyvinyl pyrrolidone (PVP); Staphylococcus aureus (S. aureus); Escherichia coli (E. coli) 1. Introduction Poly-Lactic Acid (PLA) which belongs to the family of polyglycolic acid aliphatic polyesters has received an extensive interest the last decades especially as a high-performance polymer for bio-related applications. More specific, PLA has been used in the biomedical sector for various functional objects i.e., implants [1], surgical equipment [2], nanofibrous templates for drug delivery [3], foams for tissue engineering [4], etc. PLA is a thermoplastic in nature polymeric material known also for its biocompatibility, biodegradability, superior mechanical strength compared to other thermoplastics, Biomimetics 2020, 5, 42; doi:10.3390/biomimetics5030042 www.mdpi.com/journal/biomimetics Biomimetics 2020, 5, 42 2 of 22 and ease of processing via solution and melt processing methods [5]. Moreover, for biomedical applications as for instance protective personal equipment (PPE), surgical equipment, etc., PLA offers the unique property that it can be sterilized due to its relatively high melting point (typical Tm of PLA ~150–160 ◦C), which is a prerequisite so that PLA based 3D objects can be reusable till the end of their lifetime. 3D printing is a technology in which parts are produced with the sequential addition of material layers. It belongs to the family of the additive manufacturing technologies and has received extensive scientific interest over the years. Additionally, 3D printing is in the forefront amongst other additive manufacturing technologies for on demand product development. In addition to that, 3D printing offers the unique possibility of producing 3D bulk objects consisting of different materials with various physical and chemical properties [6,7]. For these and several other practical reasons, 3D printing and additive manufacturing in general have been adopted with fervor by industry and are increasingly used as production processes. Various techniques are available nowadays for 3D printing solid materials, including Fused Filament Fabrication (FFF), direct metal laser sintering and electron beam fabrication [8]. 3D printing has been employed recently to manufacture surgical equipment [8,9], implants [10], tissue scaffolds [11], 3D bioelectronics [12], etc. One of the most promising applications of 3D printing and especially FFF technology is amongst others in the medical field for the design and development of medical devices and instruments [13,14]. Moreover, surgeons have used patient-specific computed tomography derived 3D prints for the better preoperative planning and the proper design of the surgical approach in complex operations [15–17]. In the same philosophy, 3D printed models have been used also for educational proposes of young surgeons [18,19]. Although the mechanical properties of the 3D printed have been extensively studied in literature [20], there is, however, scant literature to date for the production of multi-functional 3D printed objects, i.e., with enhanced mechanical properties [20] and additionally electrical conductivity [21], thermoelectric property [22], anti-adhesive and antimicrobial properties to avoid biofilm formation [23], etc., utilizing novel and functional materials. Recently, a great scientific focus has been devoted to the development of bulk materials with antimicrobial (AM) properties, specifically for various applications in the health sector so as to increase the environment’s hygiene and avoid the transmission and infections caused by pathogenic microorganisms [24]. Polymers as a big family of plastic materials of which many bulk 3D objects are consisting of in applications such as hospital tables, protective equipment, paints, biomedical products like implants, bandages, catheters, surgery equipment, etc. have been modified by antimicrobial agents in their bulk structure via melt-mixing or solvent mixing methods. Moreover, another promising approach to endow AM properties to bulk 3D plastic components is by depositing AM films onto their outer surface by: (i) either wet chemical methods (dip-coating using AgNP dispersions [25], sonochemical immobilization [26,27], etc.) or (ii) vacuum deposition techniques (magnetron sputtering [28], ion-beam-assisted deposition process [29], etc.). To that end, a great number of AM agents as for instance copper nanoparticles (CuNPs) and/or copper complexes, silver NPs (AgNPs) and Ag metal salts, polyhexamethylene biguanides, triclosan and chitosan based biopolymers, quaternary ammonium compounds, etc. have been reported and proven to prevent the growth of pathogenic microorganisms, such as bacteria, fungi, algae, etc. [30–32] upon being incorporated/blended into the polymer 3D bulk structure or deposited as thin films [33,34]. Amongst others, AgNPs have been in the forefront of research and several times reported for their antibacterial activity, especially due to their broad spectrum of antibacterial activity while at the same time having proven low levels of toxicity against mammalian cells [35]. However, their antimicrobial activity has not yet been fully understood. Relatively, one of the most plausible mechanisms is initially the interaction of AgNPs with the microorganism’s surface that results further to the penetration of AgNPs and/or Ag released ions (Ag+) through the cell walls, allowing them to react with the thiol group of proteins and ending up with the cell’s distortion and death [36]. Silver in its ionized form is highly reactive, as it binds to tissue proteins and endows structural changes to the bacterial cell Biomimetics 2020, 5, 42 3 of 22 wall and nuclear membrane, leading thus to cell lysis and death [37]. A side reaction of AgNPs concomitantly to their interaction with bacteria cells and bactericidal mechanism, is unavoidably the direct interaction with human cells, which leads to cytotoxicity and genotoxicity reported effects [38]. As such, it is of crucial importance AgNPs to be stabilized and/or embedded in the bulk structure of 3D component materials. Therefore, polymer/AgNP nanocomposites can be promising antimicrobial materials, which can be easily processed, i.e., via melt mixing and extrusion processes. Furthermore, polymer/AgNP nanocomposites can be extruded in the form of filaments that can be utilized further to 3D print complex and on-demand antimicrobial 3D printed bulk parts through FFF additive manufacturing technology. In relation to that, the incorporation of AgNPs in a polymer matrix by solvent mixing has been already reported [39]. The production of polymer nanocomposites by well-established industrial and large-scale processing methods as for instance melt compounding using an extruder is very advantageous [40–43]. A very promising approach to fabricate polymer/AgNP nanocomposites is via reactive melt mixing, since preformed AgNPs found as powders tend to agglomerate. The aggregation phenomena are known further to affect the material’s properties, i.e., their antimicrobial efficacy in since the interaction between the bacterial